PROJECT SUMMARY/ABSTRACT Despite significant therapeutic advances in other cancers, treatment of pancreatic ductal adenocarcinoma (PDAC) has improved only modestly over the past several decades. Consequently, pancreatic cancer remains the fourth leading cause of cancer death in the United States and a major cause of morbidity and mortality worldwide. Mutation of the proto-oncogene KRAS is a hallmark of PDAC, occurring in >90% of cases. Studies in animal models and cell lines have suggested that mutant KRAS is important for both the initiation and maintenance of PDAC, making KRAS an attractive target for therapy. Unfortunately, efforts to directly target KRAS have been thwarted by several unique features of the mutant protein, making KRAS ?undruggable? to date. Furthermore, current strategies to inhibit putative downstream signaling pathways have been largely unsuccessful. Given the high prevalence of KRAS mutations in not only PDAC but also cancers of the lung and colon, the development of new approaches to target KRAS mutant cancers remains a significant unmet need. The long-term goal of the proposed research is to improve our understanding of KRAS biology to facilitate the development of novel therapeutic strategies in PDAC and other KRAS mutant cancers. Our preliminary studies revealed that partial or complete inhibition of mutant KRAS induced marked and reversible changes in gene expression, cell signaling, and drug sensitivity, suggesting that KRAS fosters a specific cell state, potentially amenable to targeted inhibition. In this proposal, we leverage novel PDAC cell lines in which KRAS has been inactivated by CRISPR/Cas-mediated genome editing, a lentiviral cDNA library permitting expression of a diverse array of KRAS mutant variants, and CRISPR-based genome-wide screening libraries to interrogate key KRAS-relevant functional pathways and elucidate synthetic lethal interactions with KRAS mutation. Specifically, we propose to re-express different KRAS mutant variants to identify variant-specific effectors through transcriptional and biochemical profiling, offering an unprecedented evaluation of KRAS mutant variants in cancer cells. In parallel, we plan to elucidate unique genetic dependencies in cells harboring or lacking KRAS function using CRISPR-based screening. Together, these approaches are likely to reveal novel targetable pathways and proteins in KRAS mutant cells, which we will validate in additional cell lines in vitro and transplant and autochthonous models in vivo. The proposed research strategy is part of an extensive training program designed to facilitate my transition to becoming an independent investigator in basic and translational cancer biology. The main experiments will be conducted in the Jacks laboratory at the Koch Institute for Integrative Cancer Research at MIT under the guidance of Dr. Tyler Jacks, an international leader in KRAS biology and mouse models of cancer. Co- mentorship on clinical and translational science will be provided by Dr. Charles Fuchs, a leading clinical researcher and Director of the GI SPORE at the Dana-Farber Cancer Institute and Harvard Cancer Center (DF/HCC). The scientific community, encompassing the DF/HCC, Koch Institute at MIT, and Harvard Medical School, offers an outstanding environment for scientific discussion and collaboration. In addition, I have assembled a scientific advisory committee, comprised of Drs. William Hahn, Matthew Vander Heiden, and Alec Kimmelman, who collectively have a wealth of information and expertise in basic and translational research. In addition to presenting my work locally at seminars within the Harvard and MIT community, I have also proposed a comprehensive set of didactic coursework through the Harvard Catalyst program to enhance my understanding of translational research with a focus on target identification, preclinical validation, and T1 translation. As a medical oncologist and physician-scientist, I am committed to improving the care of cancer patients, with a focus on pancreatic cancer and other gastrointestinal malignancies. As a postdoctoral fellow in the Jacks lab, I have led the development of several projects, building novel tools and reagents that I will utilize in the proposed experiments. Results from these studies will form the basis for developing an independently-funded laboratory. While I have been successful in acquiring fellowship funding to support my preliminary studies, the K08 award will provide critical funding for continued mentored research and transition to independence.